Fibrinogen is a plasma protein present in blood plasma, and is a glycoprotein with a molecular weight of 340 kDa. Fibrinogen has a hexameric molecular structure wherein three chains, that is, the Aα chain, Bβ chain and γ chain, are bound to one another via disulfide bonds and the resulting complex forms a dimer (Aα-Bβ-γ)2. The Aα chain, with a size of 67 kDa, is composed of 610 amino acid residues and does not have a sugar chain. The Bβ chain, with a size of 56 kDa, is composed of 461 amino acid residues and has a sugar chain at Asn at position 364. The γ chain, with a size of 48 kDa, is composed of 411 amino acid residues and has a sugar chain at Asn at position 52.
Fibrinogen is involved in blood coagulation. In the living body, thrombin cleaves the Aα and Bβ chains of fibrinogen to remove the fibrinopeptide A and the fibrinopeptide B, thereby converting fibrinogen into (α-β-γ)2 (fibrinomonomer). The fibrinomonomer is polymerized in the presence of Ca2+ to form a fibrin polymer. Further, upon activation of blood coagulation factor XIII by thrombin, the transglutaminase activity of the factor causes formation of peptide bonds among fibrin polymers, to form strongly cross-linked fibrin.
The fibrinogen currently used in the medical field is manufactured by separation and purification from human blood plasma. Since human blood plasma is used as a raw material, there is the risk of contamination with viruses, so that a process such as inactivation of sources of infection is indispensable. Furthermore, since supply of human plasma as a raw material is dependent on blood donation, its constant and stable supply is not always easy.
By production of fibrinogen using gene recombination technology, safe fibrinogen can be stably provided. Such attempts have been reported several times so far, but none of them succeeded in providing a sufficiently efficient and satisfactory method. For example, in Patent Document 1 and Non-patent Document 1, a method for producing a recombinant fibrinogen using a Pichia yeast has been disclosed, but, in this method, fibrinogen secreted into the culture liquid is degraded by protease, so that the production method is not efficient. Patent Documents 2 and 3 disclose a method for producing a recombinant fibrinogen using animal cultured cells. However, the production is too costly and laborious to realize commercial production of fibrinogen since, for example, large-scale facilities for cell culture and control of the culture density are required. Further, since animal cells are used, there is the risk of contamination with animal-derived substances and infectious viruses, so that the method has the same problem as the current method wherein fibrinogen is manufactured from human blood.